The overall objective of this proposal is to understand the role of the endocytosis and metabolism of insulin-receptor complexes in insulin action. Further understanding in this area is essential for the eventual elucidation of the pathophysiology of insulin resistant states such as type II diabetes mellitus and obesity. Insulin initiates all of its diverse biologic actions by first binding to the high affinity insulin receptor, a glycoprotein macromolecule on the surface of target cells. This process activates the receptor as a kinase towards other substrates and also rapidly induces the internalization of the bound insulin- receptor complexes. At present, the role of the internalization process in insulin action remains poorly understood and the objective of this project is to contribute to further understanding in this important area of investigation. Accordingly, the following specific aims will be pursued: (1) To identify the structural domain of human insulin receptor which mediates its internalization and processing; (2) To generate receptor mutants defective in internalization but retaining kinase function; and (3) using such mutants, to determine the functional role of internalization of insulin-receptor complexes in insulin action on metabolic processes. The insulin-stimulated metabolic functions to be examined will include: stimulation of glucose uptake (a rapid transmembrane effect); stimulation of glycogen and protein synthesis (effects of intermediate duration); and stimulation of RNA and DNA synthesis (longer-term effects). In pursuing these aims, extensive cellular, biochemical, and molecular biological methodologies will be utilized. In brief, these will include cell culture, receptor photoaffinity labeling/processing, biosynthetic labeling, generation of receptor mutants, and measurements of several insulin-stimulated biologic functions. It is hoped that he results of the proposed studies will lead to a better understanding of the mechanism of insulin action and thereby contribute to the eventual elucidation of the pathophysiology of insulin resistance, a major abnormality in obesity and type II diabetes mellitus.